产胞外多糖酵母菌株的筛选鉴定及发酵产糖

Screening and identification of exopolysaccharide-producing yeasts

摘要:

摘要：【目的】微生物胞外多糖大多具有良好的功能和特性，但对酵母胞外多糖的研究甚少。本研究从自然界中筛选出产胞外多糖的酵母菌株，并对其发酵产糖条件进行初步研究。【方法】利用平板涂布法从自然界中分离得到酵母菌株，苯酚硫酸法测定菌株胞外多糖的产量，筛选出胞外多糖高产菌株，并对其进行5.8S rDNA分类鉴定，最后优化其产糖培养基组成。【结果】对从葡萄、蜜枣、土壤样品中分离得到的132株酵母进行筛选，最终得到3株高产胞外多糖的酵母菌株Z14、Z20和L25。经5.8S rDNA序列测定及系统发育分析，从左优红葡萄中分离得到的Z14和Z20与东方伊萨酵母（Issatchenkia orientalis）处于同一分支，相似性达到99 %以上；从落叶松近表层土壤分离得到的L25与土生隐球酵母（Cryptococcus humicolus）处于同一分支，相似性为98.8 %。经优化，利于Z20胞外多糖合成的最优发酵培养基配方为：葡萄糖8 %，(NH4)SO4 0.2 %，KH2PO4 0.1 %，酵母浸粉0.1 %，CaCl2 0.01 %。在初始pH 6.0，发酵温度28 ℃，摇床转数160 r/min条件下，在此培养基中发酵4 d后胞外多糖产量可达2.046 g/L，比复筛时的产量1.137 g/L提高了79.9 %。【结论】文献已报道某些属的酵母可以生产胞外多糖，经本文研究发现Issatchenkia属的酵母也可以合成胞外多糖，并且改变基础产糖培养基的成分可以显着提高Z20胞外多糖的产量。

Abstract:

Abstract: [Objective] Most of microbial extracellular polysaccharide (EPS) has the favorable functionality and peculiarity, but little research has been done about EPS that were synthesized by yeasts. Screening of EPS-producing yeasts and studies on the optimal culture composition are the main purpose of this study. [Methods] Yeasts were obtained by screening from natural samples using spread-plate method, Yield of EPS synthesized by yeasts was estimated by phenol-sulfate acid method, Based on 5.8S rDNA sequence determination, three strains were identified and selected for the optimization of fermentative medium. [Results] 132 yeast strains were isolated from grapes, preserves and topsoil. Three yeast strains were proved to be as high exopolysaccharide-producing yeasts. The results of 5.8S rDNA sequence determination showed that similarities of two yeasts isolated from “Zuoyouhong” grape (Z14 and Z20) were over 99 % compared with Issatchenkia orientalis, meanwhile another yeast L25 isolated from topsoil of Larix gmelini was identified as Cryptococcus humicolus, with the similarity of 98.8 %. The optimal medium for strain Z20 for producing EPS was as followed: 8 % glucose, 0.2 % (NH4)SO4 , 0.1 % KH2PO4 , 0.1 % yeast extract powder and 0.01 %CaCl2 . Under the condition of initial pH 6.0, 28 ℃ and 160 r/min for 4 d, yield of EPS could reach to 2.046 g/L, which was 79.9 % higher than the control (1.137 g/L). [Conclusion] Reports that yeasts belong to some genera possessed the abilities to synthesize EPS could be seen in some related references, according to the study, we could get the conclusion that yeasts belong to Issatchenkia can synthesize EPS and EPS yield of stain Z20 could be increased by changing culture composition.